Feature Articles

Cell Culture Central to Upstream Scaleup

Why There's a Good Deal of Truth to the Dictum: "As Cells Go, So Goeth the Process"

Scaleup trends closely follow the larger bioprocessing industry. According to Christian Kaisermayer, Ph.D., project manager for cell culture at GE Healthcare Life Sciences the two significant drivers are process flexibility and complete solutions.

Underlying both trends are the rise in protein titers over the past decade and disposable bioprocessing.

Disposables play an obvious role in scaleup, i.e., that form factors for smaller single-use cell culture vessels (e.g. GE Healthcare’s Wave Bioreactor™ systems) can be identical to production-scale equipment, thus facilitating scaledown exercises. GE offers Wave systems as “segmented” bags for parallel experiments or screening, with working volumes in each compartment ranging in volume from 50–250 mL. Each chamber is controlled for temperature and headspace gas composition.

“The upstream capacity crunch discussed just a few years ago never occurred,” Dr. Kaisermayer says. “Accordingly, most products in development today will never see 10,000 to 20,000 liter bioreactors. Thus the focus on flexibility. Moreover, the sweet spot transition volume between disposable and stainless steel reactors has already surpassed 1,000 liters.”

With few exceptions today’s production engineers consider disposables at every level of process design, from benchtop to pilot scale and beyond. The more rapid setup and reduced cleaning and related validation associated with single-use bioprocessing improve equipment utilization—not just for final production but also during development.

“Personnel can now concentrate on their experiments rather than spending time on cleaning and sterilization,” explains Dr. Kaisermayer.

On the subject of integration and “complete solutions,” GE Healthcare’s ReadyToProcess™ product line covers all unit operations from media preparation and cell cultivation to protein purification. Customers can source stand-alone or complete single-use technologies for end-to-end processes from GE and even engage their Enterprise Solutions team to design, develop, and execute entire ready-to-use bioproduction facilities.

Even in situations where development occurs in plastic, and production in stainless steel, the disconnect need not be problematic.

“Transferring a process from disposables to stainless can be controversial in terms of reactor engineering and fluid dynamics,” says Dr. Kaisermayer. “Some experts feel that cells should be shaken, others that cells should be stirred. It usually doesn’t matter because agitation intensity can be adapted to the robustness of the cell line.”

No Impellers Needed

PBS Biotech designs and manufactures single-use bioreactors with a twist. According to Daniel Giroux, vp of R&D, duplicating the agitation mechanisms of small reactors in large systems is problematic without subjecting cells to significant shear damage.

PBS’ U-shaped bioreactors employ a patented mixing technology, the Air-Wheel™, that exploits the fact that cell cultures must be oxygenated. Air-Wheel uses the buoyancy of rising gas bubbles to turn the Air-Wheel structure, thus avoiding the need for external actuation of the stirring mechanism.

The result, according to the company, is rapid, homogeneous mixing and a high mass transfer rate with extremely low shear stress on cells.

PBS offers scalability from three liters for lab work, and 80 liters for preclinical and clinical batches. In targeting full-scale GMP production, the company plans to produce a 500 L system later this year, and a 2,500 L bioreactor in 2012.

“Since the mixing mechanism is proportional to the size of the vessel, Air-Wheel works more efficiently as bioreactor volume increases,” explains Giroux, a definite benefit given that mixing is one of scaleup’s biggest challenges.

According to Giroux, traditional bioreactors were designed for steam sterilization—essentially as pressure vessels. “They were never optimized for mixing. Mammalian cell culture reactors were adapted from chemical mixing tanks, which used shaft-and-impeller mixing.”

As bioreactors grew in size, designers were more or less forced to retain the original form factor and stirring mechanism, including impeller design. “Which is why mixing and mass-transfer problems get worse as the size increases.”

Designers of traditional single-use reactors, he says, never fully appreciated that cleaning was unnecessary. Except for Wave, now part of GE, they mostly retained the shape and mechanical characteristics of traditional tanks. “But it becomes difficult to put impellers into the right locations.”

He claims that at three liters, glass or stainless steel reactors exhibit up to ten times as much shear as the PBS bioreactors. “Our shear goes up a bit at higher scale, but very little.” Ditto for mixing time. Time to 95% mixing is claimed to be less than one minute.

“If you have a tool where critical parameters are so similar across all scales, scaleup will be much easier.”

What about cells for which shear is not a problem? Many such lines exist, Giroux says, but for early-stage development lines, where shear sensitivity is unknown, knowing that shear will not be a problem across production scales is no small comfort.

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